Recent advances in recombinant DNA technology have brought the substantial promise of gene therapy nearer to realization. Techniques for specifically altering the nucleotide sequence of a DNA molecule have been a particularly valuable development. These techniques enable a DNA molecule to be tailored to provide a specific selected effect, when provided as gene therapy.
An alternative to gene therapy is to manipulate gene expression at the RNA level. Targeting RNA has several advantages not inherent in gene manipulation at the DNA level. For example, RNA therapy can be targeted to cells and tissues where gene expression is actually occurring, i.e., cells in which mRNA encoding a specific gene product is being actively transcribed and translated. Additionally, in certain situations, RNA is a good target for gene therapy because RNA is an intermediate in reverse transcription, whereby certain viral DNA genomes are produced.
Already, RNA-targeted gene therapy has been accomplished through the use of anti-sense RNA molecules (RNA or DNA) designed specifically to block the translation of a target RNA molecule into protein. Assuming knowledge of the nature of the target mRNA molecule, complementary oligonucleotides are created to bind specifically to regions of the RNA molecule critical for selected functions of the molecule, such as maintenance of secondary structure or translation, thereby disrupting expression of the gene product encoded by the RNA.
Another type of RNA-targeted gene therapy involves the use of sequence-specific endoribonucleases, known as ribozymes, to achieve cleavage and inactivation of gene transcripts in vivo. According to this strategy, assuming knowledge of the mRNA transcribed by the gene, ribozymes can be synthesized which hybridize specifically to a predetermined sequence and cleave the RNA molecule at a specified site in the target sequence, so as to inhibit production of the gene product by destroying its messenger.
Both ribozymes and anti-sense molecules are capable of disrupting gene expression at the RNA level, ribozymes by targeted destruction of the specific RNA, and anti-sense molecules by disrupting secondary structure or blocking translation.
Although inhibiting gene expression is certainly a valuable mode of RNA therapy, it would be even better to have RNA-targeted methods for enhancing gene expression (by increasing the stability or translation efficiency of an RNA molecule) or altering the subsequently produced gene product. Such methods are available for DNA-targeted gene therapy (e.g., in vitro mutagenesis), but not for RNA-targeted gene therapy. Clearly, a method for altering the nucleic acid sequence of an RNA molecule could provide many new therapeutic methods by combining the advantages inherent in specifically altering DNA with the advantages of targeting RNA.